9 results on '"Tetsuo Ushiku"'
Search Results
2. N-terminal peptide fragment constitutes core of amyloid deposition of serum amyloid A: An imaging mass spectrometry study
- Author
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Yukako Shintani-Domoto, Yuki Sugiura, Makiko Ogawa, Eiji Sugiyama, Hiroyuki Abe, Takashi Sakatani, Ryuji Ohashi, Tetsuo Ushiku, and Masashi Fukayama
- Subjects
Medicine ,Science - Abstract
Serum amyloid A (SAA) is an acute phase protein, which undergoes structural changes and deposits in the extracellular matrix, causing organ damage. Systemic AA amyloidosis is a relatively common amyloid subtype among the more than 30 amyloid subtypes, but the mechanism of amyloid fibril formation remains unclear. In this study, we investigated the tissue distribution of SAA derived peptides in formalin-fixed paraffin embedded (FFPE) specimens of human myocardium with amyloidosis using matrix-assisted laser desorption/ionization imaging mass spectrometry (MALDI-IMS). In the whole SAA protein, four trypsin-digested peptides in the range of SAA2-67 were visualized and the N-terminal peptide; SAA2-15, was selectively localized in the Congo red-positive region. The C-terminal peptides; SAA47-62, SAA48-62, and SAA63-67 were detected not only in the Congo red-positive region but also in the surrounding negative region. Our results demonstrate that the N-terminal SAA2-15 plays a critical role in the formation of AA amyloid fibril, as previously reported. Roles of the C-terminal peptides require further investigation.
- Published
- 2022
3. Virus-host interactions in carcinogenesis of Epstein-Barr virus-associated gastric carcinoma: Potential roles of lost ARID1A expression in its early stage.
- Author
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Hiroyuki Abe, Akiko Kunita, Yuya Otake, Teru Kanda, Atsushi Kaneda, Tetsuo Ushiku, and Masashi Fukayama
- Subjects
Medicine ,Science - Abstract
Epstein-Barr virus (EBV)-associated gastric carcinoma (EBVaGC) is a distinct molecular subtype of gastric cancer characterized by viral infection and cellular abnormalities, including loss of AT-rich interaction domain 1A (ARID1A) expression (lost ARID1A). To evaluate the significance of lost ARID1A in the development of EBVaGC, we performed in situ hybridization of EBV-encoded RNA (EBER) and immunohistochemistry of ARID1A in the non-neoplastic gastric mucosa and intramucosal cancer tissue of EBVaGC with in vitro infection analysis of ARID1A-knockdown and -knockout gastric cells. Screening of EBER by in situ hybridization revealed a frequency of approximately 0.2% EBER-positive epithelial cells in non-neoplastic gastric mucosa tissue samples. Six small foci of EBV-infected epithelial cells showed two types of histology: degenerated (n = 3) and metaplastic (n = 3) epithelial cells. ARID1A was lost in the former type. In intramucosal EBVaGC, there were ARID1A-lost (n = 5) and -preserved tumors (n = 7), suggesting that ARID1A-lost carcinomas are derived from ARID1A-lost precursor cells in the non-neoplastic mucosa. Lost ARID1A was also observed in non-neoplastic mucosa adjacent to an ARID1A-lost EBVaGC. In vitro experiments using siRNA knockdown and the CRISPR/Cas9-knockout system demonstrated that transient reduction or permanent loss of ARID1A expression markedly increased the efficiency of EBV infection to stomach epithelial cells. Taken together, lost ARID1A plays a role in initiating EBV-driven carcinogenesis in stomach epithelial cells, which develop to a distinct subtype of EBVaGC within the proper mucosal layer. Lost ARID1A is one of the constituents of virus-host interactions in the carcinogenesis of EBVaGC.
- Published
- 2021
- Full Text
- View/download PDF
4. Mass spectrometry-based absolute quantification of amyloid proteins in pathology tissue specimens: Merits and limitations.
- Author
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Makiko Ogawa, Yukako Shintani-Domoto, Yoshiki Nagashima, Koji L Ode, Aya Sato, Yoshihiro Shimizu, Kenichi Ohashi, Michael H A Roehrl, Tetsuo Ushiku, Hiroki R Ueda, and Masashi Fukayama
- Subjects
Medicine ,Science - Abstract
To clarify the significance of quantitative analyses of amyloid proteins in clinical practice and in research relating to systemic amyloidoses, we applied mass spectrometry-based quantification by isotope-labeled cell-free products (MS-QBIC) to formalin-fixed, paraffin-embedded (FFPE) tissues. The technique was applied to amyloid tissues collected by laser microdissection of Congo red-stained lesions of FFPE specimens. Twelve of 13 amyloid precursor proteins were successfully quantified, including serum amyloid A (SAA), transthyretin (TTR), immunoglobulin kappa light chain (IGK), immunoglobulin lambda light chain (IGL), beta-2-microglobulin (B2M), apolipoprotein (Apo) A1, Apo A4, Apo E, lysozyme, Apo A2, gelsolin, and fibrinogen alpha chain; leukocyte cell-derived chemotaxin-2 was not detected. The quantification of SAA, TTR, IGK, IGL, and B2M confirmed the responsible proteins, even when the immunohistochemical results were not decisive. Considerable amounts of Apo A1, Apo A4, and Apo E were deposited in parallel amounts with the responsible proteins. Quantification of amyloid protein by MS-QBIC is feasible and useful for the classification of and research on systemic amyloidoses.
- Published
- 2020
- Full Text
- View/download PDF
5. Heterozygous knockout of Bile salt export pump ameliorates liver steatosis in mice fed a high-fat diet.
- Author
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Kazuya Okushin, Takeya Tsutsumi, Kazuhiko Ikeuchi, Akira Kado, Kenichiro Enooku, Hidetaka Fujinaga, Naoko Yamauchi, Tetsuo Ushiku, Kyoji Moriya, Hiroshi Yotsuyanagi, and Kazuhiko Koike
- Subjects
Medicine ,Science - Abstract
The incidence of nonalcoholic steatohepatitis (NASH) is increasing worldwide, including in Asian countries. We reported that the hepatic expression of bile salt export pump (BSEP) was downregulated in patients with NASH, suggesting that BSEP is involved in the pathogenesis of NASH. To identify the underlying mechanism, we analyzed Bsep heterozygous knock-out (Bsep+/- mice) and wild-type (WT) C57BL/6J mice fed a high-fat diet (HFD) (32.0% animal fat) or normal diet. We examined histological changes, levels of hepatic lipids and hepatic bile acids, and expression of genes related to bile acid and cholesterol metabolism. HFD-fed Bsep+/- mice exhibited milder hepatic steatosis and less weight gain, compared to HFD-fed WT mice. The concentrations of total bile acid, triglycerides, and cholesterols were reduced in the liver of HFD-fed Bsep+/- mice. Regarding hepatic bile acid metabolism, the expression levels of Farnesoid X receptor (Fxr) and Multidrug resistance-associated protein 2 were significantly upregulated in HFD-fed Bsep+/- mice, compared to HFD-fed WT mice. Furthermore, several alterations were observed in upstream cholesterol metabolism in the liver. The expression levels of bile acid metabolism-related genes were also altered in the intestine of HFD-fed Bsep+/- mice. In conclusion, HFD-fed Bsep+/- mice exhibited significant alterations of the expression levels of genes related to bile acid and lipid metabolism in both the liver and ileum, resulting in alleviated steatosis and less weight gain. These results suggest the importance of BSEP for maintenance of bile acid and cholesterol metabolism. Further investigations of the involvement of BSEP in the pathogenesis of NASH will provide greater insight and facilitate the development of novel therapeutic modalities.
- Published
- 2020
- Full Text
- View/download PDF
6. Tumor Content Chart-Assisted HER2/CEP17 Digital PCR Analysis of Gastric Cancer Biopsy Specimens.
- Author
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Keisuke Matsusaka, Shumpei Ishikawa, Atsuhito Nakayama, Tetsuo Ushiku, Aiko Nishimoto, Masayuki Urabe, Nobuyuki Kaneko, Akiko Kunita, Atsushi Kaneda, Hiroyuki Aburatani, Mitsuhiro Fujishiro, Yasuyuki Seto, and Masashi Fukayama
- Subjects
Medicine ,Science - Abstract
Evaluating HER2 gene amplification is an essential component of therapeutic decision-making for advanced or metastatic gastric cancer. A simple method that is applicable to small, formalin-fixed, paraffin-embedded biopsy specimens is desirable as an adjunct to or as a substitute for currently used HER2 immunohistochemistry and in situ hybridization protocols. In this study, we developed a microfluidics-based digital PCR method for determining HER2 and chromosome 17 centromere (CEP17) copy numbers and estimating tumor content ratio (TCR). The HER2/CEP17 ratio is determined by three variables-TCR and absolute copy numbers of HER2 and CEP17-by examining tumor cells; only the ratio of the latter two can be obtained by digital PCR using the whole specimen without purifying tumor cells. TCR was determined by semi-automatic image analysis. We developed a Tumor Content chart, which is a plane of rectangular coordinates consisting of HER2/CEP17 digital PCR data and TCR that delineates amplified, non-amplified, and equivocal areas. By applying this method, 44 clinical gastric cancer biopsy samples were classified as amplified (n = 13), non-amplified (n = 25), or equivocal (n = 6). By comparison, 11 samples were positive, 11 were negative, and 22 were equivocally immunohistochemistry. Thus, our novel method reduced the number of equivocal samples from 22 to 6, thereby obviating the need for confirmation by fluorescence or dual-probe in situ hybridization to < 30% of cases. Tumor content chart-assisted digital PCR analysis is also applicable to multiple sites in surgically resected tissues. These results indicate that this analysis is a useful alternative to HER2 immunohistochemistry in gastric cancers that can serve as a basis for the automated evaluation of HER2 status.
- Published
- 2016
- Full Text
- View/download PDF
7. Mass spectrometry-based absolute quantification of amyloid proteins in pathology tissue specimens: Merits and limitations
- Author
-
Yoshihiro Shimizu, Tetsuo Ushiku, Makiko Ogawa, Koji L. Ode, Kenichi Ohashi, Aya Sato, Yoshiki Nagashima, Hiroki R. Ueda, Masashi Fukayama, Yukako Shintani-Domoto, and Michael H.A. Roehrl
- Subjects
0301 basic medicine ,Male ,Apolipoprotein B ,Light ,Physiology ,Immunostaining ,Pathology and Laboratory Medicine ,Biochemistry ,Mass Spectrometry ,0302 clinical medicine ,Immune Physiology ,Medicine and Health Sciences ,Post-Translational Modification ,Laser capture microdissection ,Fibrinogen alpha chain ,Staining ,Aged, 80 and over ,Multidisciplinary ,Immune System Proteins ,biology ,Chemistry ,Amyloidosis ,Physics ,Electromagnetic Radiation ,Middle Aged ,Immunohistochemistry ,Enzymes ,030220 oncology & carcinogenesis ,Physical Sciences ,Medicine ,Female ,Signal Peptides ,Research Article ,Amyloid ,Science ,Lipoproteins ,Immunology ,Lysozyme ,Amyloidogenic Proteins ,Research and Analysis Methods ,Antibodies ,03 medical and health sciences ,Signs and Symptoms ,Diagnostic Medicine ,medicine ,Humans ,Serum amyloid A ,Aged ,Biology and Life Sciences ,Proteins ,medicine.disease ,Molecular biology ,Transthyretin ,030104 developmental biology ,Apolipoproteins ,Specimen Preparation and Treatment ,biology.protein ,Amyloid Proteins ,Enzymology ,Gelsolin - Abstract
To clarify the significance of quantitative analyses of amyloid proteins in clinical practice and in research relating to systemic amyloidoses, we applied mass spectrometry-based quantification by isotope-labeled cell-free products (MS-QBIC) to formalin-fixed, paraffin-embedded (FFPE) tissues. The technique was applied to amyloid tissues collected by laser microdissection of Congo red-stained lesions of FFPE specimens. Twelve of 13 amyloid precursor proteins were successfully quantified, including serum amyloid A (SAA), transthyretin (TTR), immunoglobulin kappa light chain (IGK), immunoglobulin lambda light chain (IGL), beta-2-microglobulin (B2M), apolipoprotein (Apo) A1, Apo A4, Apo E, lysozyme, Apo A2, gelsolin, and fibrinogen alpha chain; leukocyte cell-derived chemotaxin-2 was not detected. The quantification of SAA, TTR, IGK, IGL, and B2M confirmed the responsible proteins, even when the immunohistochemical results were not decisive. Considerable amounts of Apo A1, Apo A4, and Apo E were deposited in parallel amounts with the responsible proteins. Quantification of amyloid protein by MS-QBIC is feasible and useful for the classification of and research on systemic amyloidoses.
- Published
- 2020
8. Heterozygous knockout of Bile salt export pump ameliorates liver steatosis in mice fed a high-fat diet
- Author
-
Takeya Tsutsumi, Hidetaka Fujinaga, Kazuhiko Ikeuchi, Hiroshi Yotsuyanagi, Naoko Yamauchi, Kazuya Okushin, Tetsuo Ushiku, Kyoji Moriya, Kenichiro Enooku, Akira Kado, and Kazuhiko Koike
- Subjects
Male ,Steatosis ,Physiology ,Gene Expression ,Nonalcoholic Steatohepatitis ,Pathology and Laboratory Medicine ,Biochemistry ,Cytopathology ,chemistry.chemical_compound ,Mice ,Non-alcoholic Fatty Liver Disease ,Medicine and Health Sciences ,Bile ,ATP Binding Cassette Transporter, Subfamily B, Member 11 ,Mice, Knockout ,Multidisciplinary ,Bile acid ,Liver Diseases ,Fatty liver ,Animal Models ,Lipids ,Body Fluids ,Cholesterol ,Liver ,Experimental Organism Systems ,Medicine ,Anatomy ,Research Article ,medicine.medical_specialty ,Heterozygote ,Normal diet ,medicine.drug_class ,Science ,Mouse Models ,Gastroenterology and Hepatology ,Diet, High-Fat ,Research and Analysis Methods ,digestive system ,Bile Acids and Salts ,Model Organisms ,Ileum ,Internal medicine ,medicine ,Genetics ,Animals ,nutritional and metabolic diseases ,Biology and Life Sciences ,Lipid metabolism ,medicine.disease ,Lipid Metabolism ,Bile Salt Export Pump ,Mice, Inbred C57BL ,Fatty Liver ,Gastrointestinal Tract ,Disease Models, Animal ,Endocrinology ,chemistry ,Anatomical Pathology ,Animal Studies ,Farnesoid X receptor ,Digestive System - Abstract
The incidence of nonalcoholic steatohepatitis (NASH) is increasing worldwide, including in Asian countries. We reported that the hepatic expression of bile salt export pump (BSEP) was downregulated in patients with NASH, suggesting that BSEP is involved in the pathogenesis of NASH. To identify the underlying mechanism, we analyzed Bsep heterozygous knock-out (Bsep+/- mice) and wild-type (WT) C57BL/6J mice fed a high-fat diet (HFD) (32.0% animal fat) or normal diet. We examined histological changes, levels of hepatic lipids and hepatic bile acids, and expression of genes related to bile acid and cholesterol metabolism. HFD-fed Bsep+/- mice exhibited milder hepatic steatosis and less weight gain, compared to HFD-fed WT mice. The concentrations of total bile acid, triglycerides, and cholesterols were reduced in the liver of HFD-fed Bsep+/- mice. Regarding hepatic bile acid metabolism, the expression levels of Farnesoid X receptor (Fxr) and Multidrug resistance-associated protein 2 were significantly upregulated in HFD-fed Bsep+/- mice, compared to HFD-fed WT mice. Furthermore, several alterations were observed in upstream cholesterol metabolism in the liver. The expression levels of bile acid metabolism-related genes were also altered in the intestine of HFD-fed Bsep+/- mice. In conclusion, HFD-fed Bsep+/- mice exhibited significant alterations of the expression levels of genes related to bile acid and lipid metabolism in both the liver and ileum, resulting in alleviated steatosis and less weight gain. These results suggest the importance of BSEP for maintenance of bile acid and cholesterol metabolism. Further investigations of the involvement of BSEP in the pathogenesis of NASH will provide greater insight and facilitate the development of novel therapeutic modalities.
- Published
- 2020
9. Tumor Content Chart-Assisted HER2/CEP17 Digital PCR Analysis of Gastric Cancer Biopsy Specimens
- Author
-
Yasuyuki Seto, Akiko Kunita, Atsuhito Nakayama, Masashi Fukayama, Masayuki Urabe, Nobuyuki Kaneko, Atsushi Kaneda, Hiroyuki Aburatani, Tetsuo Ushiku, Shumpei Ishikawa, Mitsuhiro Fujishiro, Aiko Nishimoto, and Keisuke Matsusaka
- Subjects
Male ,0301 basic medicine ,Pathology ,Tissue Fixation ,Receptor, ErbB-2 ,Biopsy ,Microfluidics ,Gene Dosage ,Gene Expression ,lcsh:Medicine ,Artificial Gene Amplification and Extension ,Polymerase Chain Reaction ,Infographics ,law.invention ,0302 clinical medicine ,Cell Signaling ,law ,Breast Tumors ,Medicine and Health Sciences ,Digital polymerase chain reaction ,lcsh:Science ,In Situ Hybridization, Fluorescence ,Polymerase chain reaction ,Multidisciplinary ,medicine.diagnostic_test ,Charts ,Immunohistochemistry ,Oncology ,Data Interpretation, Statistical ,030220 oncology & carcinogenesis ,Genomic Signal Processing ,Research Article ,Signal Transduction ,Computer and Information Sciences ,medicine.medical_specialty ,Surgical and Invasive Medical Procedures ,In situ hybridization ,Biology ,Research and Analysis Methods ,Gene dosage ,Digestive System Procedures ,03 medical and health sciences ,Breast cancer ,Stomach Neoplasms ,Cell Line, Tumor ,Formaldehyde ,Gastrointestinal Tumors ,Breast Cancer ,medicine ,Humans ,Molecular Biology Techniques ,Molecular Biology ,Automation, Laboratory ,Tissue Embedding ,Data Visualization ,lcsh:R ,Biology and Life Sciences ,Cancers and Neoplasms ,Cancer ,Cell Biology ,medicine.disease ,Gastric Cancer ,030104 developmental biology ,lcsh:Q ,Chromosomes, Human, Pair 17 - Abstract
Evaluating HER2 gene amplification is an essential component of therapeutic decision-making for advanced or metastatic gastric cancer. A simple method that is applicable to small, formalin-fixed, paraffin-embedded biopsy specimens is desirable as an adjunct to or as a substitute for currently used HER2 immunohistochemistry and in situ hybridization protocols. In this study, we developed a microfluidics-based digital PCR method for determining HER2 and chromosome 17 centromere (CEP17) copy numbers and estimating tumor content ratio (TCR). The HER2/CEP17 ratio is determined by three variables—TCR and absolute copy numbers of HER2 and CEP17—by examining tumor cells; only the ratio of the latter two can be obtained by digital PCR using the whole specimen without purifying tumor cells. TCR was determined by semi-automatic image analysis. We developed a Tumor Content chart, which is a plane of rectangular coordinates consisting of HER2/CEP17 digital PCR data and TCR that delineates amplified, non-amplified, and equivocal areas. By applying this method, 44 clinical gastric cancer biopsy samples were classified as amplified (n = 13), non-amplified (n = 25), or equivocal (n = 6). By comparison, 11 samples were positive, 11 were negative, and 22 were equivocally immunohistochemistry. Thus, our novel method reduced the number of equivocal samples from 22 to 6, thereby obviating the need for confirmation by fluorescence or dual-probe in situ hybridization to < 30% of cases. Tumor content chart-assisted digital PCR analysis is also applicable to multiple sites in surgically resected tissues. These results indicate that this analysis is a useful alternative to HER2 immunohistochemistry in gastric cancers that can serve as a basis for the automated evaluation of HER2 status.
- Published
- 2016
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